| Rheumatoid arthritis(RA)is a relatively common autoimmune disease.Its clinical manifestations are mostly morning stiffness and joint involvement.In the later stage,the disease can cause joint deformities,lingering and difficult to heal,which seriously affects the patient's normal life and work.Its pathological manifestations include chronic inflammation of the synovial membrane,accumulation of fluid in the joint cavity,and formation of pannus,which gradually causes erosion of articular cartilage and bone.The etiology and pathogenesis of the disease have not been fully clarified,and there is currently no radical cure.RA belongs to the category of "Arthralgia" in Traditional Chinese Medicine.Traditional Chinese Medicine has accumulated rich experience in the treatment of "Arthralgia".Shaogan Fuzi Decoction is derived from "Treatise on Febrile Diseases".The whole prescription is composed of three Chinese herbs:peony,roasted licorice,and aconite.This prescription and its addition and subtraction prescriptions are clinically effective in treating RA,but there is still a lack of systematic pharmacodynamics and mechanism research.This study established collageninduced arthritis(CIA)rat model to observe the effects of Shaogan Fuzi Decoction on the swelling of the foot and plantar,arthritis index,ankle joint pathology,and imaging changes in CIA rats.And through the PPAR-?/NF-?B signaling pathway,a preliminary exploration of its mechanism of action in the treatment of RA provides a reliable experimental basis for its further clinical application.1 Therapeutic effect of Shaogan Fuzi Decoction on CIA model ratsObjective:To observe the effects of Shaogan Fuzi Decoction on the swelling of the foot,arthritis index,ankle joint pathology,and imaging changes in CIA rats,to clarify the efficacy of the prescription in the treatment of RA.Methods:SPF Wistar rats,12 were randomly selected as the normal group,and the remaining rats were used bovine type ? collagen and incomplete Freund's adjuvant to prepare RA rat models.On the first day of the experiment(d 1),the emulsifier of bovine type ?collagen and incomplete Freund's adjuvant was injected intracutaneously at multiple points at the tail root of rats,0.2 mL each rat.On the 7th day(d 7),rats have injected with the emulsifier,(0.1 mL each rat)and measured for foot swelling.On the 14th day(d 14),the rats were scored according to the arthritis index scoring standard.The rats with an AI score?of 4 points were considered as successful models.They were divided into model groups,Shaogan Fuzi Decoction low-dose group(1.05 g/kg),Shaogan Fuzi Decoction medium-dose group(2.1 g/kg),Shaogan Fuzi Decoction high-dose group(4.2 g/kg),and Tripterygium glycoside tablet group(9 mg/kg),15 rats in each group.On the 15th day(d 15),each group was gavaged according to the above dose.The normal group and the model group were given the same dose of normal saline for 4 consecutive weeks.24 hours after the last administration,the rats in each group were anesthetized,and the ankle joints were taken,then HE stained,and Micro-CT scan to observe the pathological and imaging changes.Results:?Weight:Compared with the normal group,the model group had a tendency to lose weight after d 14,and the weight decreased significantly from d 21 to the end of the experiment(P<0.01).Compared with the CIA model group,Shaogan Fuzi Decoction lowdose group after two weeks and four weeks of administration,Shaogan Fuzi Decoction medium-dose group after two weeks of administration,Tripterygium glycoside tablet group after two weeks,three weeks and four weeks of administration,bodyweight significantly increased(P<0.05 or 0.01).?The left foot swelling degree:On d 14 after the initial immunization,compared with the normal group,the swelling degree of the left foot of the CIA model group was significantly increased(P<0.01).Compared with the model group,Shaogan Fuzi Decoction low-dose group decreased significantly after one week of administration(P<0.01),while the Shaogan Fuzi Decoction medium,high-dose group,and Tripterygium glycoside tablet group decreased significantly after two weeks of administration(P<0.05 or 0.01).?The right foot Swelling degree:On d 14 after the initial immunization,compared with the normal group,the swelling degree of the right hind foot of the CIA model group was significantly increased(P<0.01).Compared with the model group.the Shaogan Fuzi Decoction low-dose,medium-dose group and the Tripterygium glucoside tablet group were significantly reduced after one week of administration(P<0.05 or 0.01),while the Shaogan Fuzi Decoction high-dose group was significantly reduced after two weeks of administration(P<0.05 or 0.01).?Arthritis index score:On d 14 after the initial immunization,compared with the normal group,the arthritis index score of the rats in the CIA model group was significantly higher than that in the normal group(P<0.01).Compared with the model group.Shaogan Fuzi Decoction and Tripterygium glucoside tablet group were significantly reduced after two weeks of administration(P<0.05 or 0.01).?Ankle joint pathology:Compared with the normal group,the ankle joint pathology score of the model group was significantly increased(P<0.01).Compared with the model group,in the middle and high dose groups of Shaogan Fuzi Decoction and Tripterygium glucoside tablet group,the score of the ankle joint was significantly reduced(P<0.05 or 0.01).Compared with the normal group,the degree of synovial epithelial cell lesions in the model group was increased(P<0.01);compared with the model group,the degree of synovial epithelial cell lesions in the high-dose Shaogan Fuzi Decoction group and Tripterygium glucoside tablet group was reduced(P<0.05).Compared with the normal group,the degree of synovial stromal cell lesions in the model group was increased(P<0.01);compared with the model group,the degree of synovial stromal cell lesions in each dose group of Shaogan Fuzi Decoction and Tripterygium glycoside tablet group were reduced(P<0.05 or 0.01).Compared with the normal group,the degree of synovial cartilage lesions in the model group was increased(P<0.01);compared with the model group,the degree of the synovial cartilage tissue of the Shaogan Fuzi Decoction group and the Tripterygium glucoside tablet group were reduced(P<0.05 or 0.01).?Micro-CT imaging:Micro-CT scan showed that each dose group of Shaogan Fuzi Decoction and Tripterygium glucoside tablet group could ameliorate joint swelling,joint synovial hyperplasia,and bone destruction in CIA rats.Conclusion:Shaogan Fuzi Decoction can reduce the foot swelling in CIA model rats,decrease the arthritis index score,ameliorate the pathological damage of the ankle joint,and alleviate the synovial hyperplasia.This prescription has a certain therapeutic effect on the CIA model rats.2 Mechanism research of Shaogan Fuzi Decoction on intervention of CIA ratsObjective:To observe the effect of Shaogan Fuzi Decoction on the contents of PGE2,NO,TNF-?,IL-1?,IL-6 in serum and PPAR-y,NF-?B p65,p-NF-?B p65,COX-2,iNOS protein expression in ankle joint of CIA model rats,to research the mechanism of Shaogan Fuzi Decoction in the treatment of RA based on PPAR-?/NF-?B signaling pathway.Methods:The CIA rat model of rheumatoid arthritis was prepared with bovine type ?collagen and incomplete Freund's adjuvant.Arthritis index score was performed on the 14th day(d 14)of the experiment.Rats with a score of?4 were considered to be successful and were divided into the Shaogan Fuzi Decoction low-dose group(1.05 g/kg),Shaogan Fuzi Decoction medium-dose group(2.1 g/kg),Shaogan Fuzi Decoction high dose group(4.2 g/kg),and tripterygium glucoside tablet group(9 mg/kg).On the 15th day of the experiment(d 15),each group was gavaged according to the dose.The normal group and the model group were given the same dose of normal saline for 4 consecutive weeks.The rats were anesthetized 24 hours after the last administration,and serum,bilateral knee joint synovium,and ankle joints were collected.Colorimetric method to detect serum NO level,ELISA method to detect serum PGE2,TNF-?,IL-1?,and IL-6 level,and immunohistochemical method to detect PPAR-?,NF?B p65,p-NF-?B p65,COX-2,and iNOS protein expression in the ankle joint.Results:Compared with the normal group,the serum levels of PGE2,NO,TNF-?,IL-1?,and IL-6 increased significantly in the CIA model group(P<0.01).Compared with the model group,the PGE2 content of Shaogan Fuzi Decoction low,medium and high-dose groups decreased(P<0.01),and the NO content of Shaogan Fuzi Decoction low-dose group and Tripterygium glucoside tablet group decreased(P<0.05).The levels of TNF-? in the low and medium dose groups of Shaogan Fuzi Decoction and the Tripterygium glucoside tablet group were significantly decreased(P<0.05 or 0.01),and the levels of IL-1? and IL-6 in the low and medium dose groups of Shaogan Fuzi Decoction decreased significantly compared with the model group(P<0.05).Compared with the normal group,the protein expression of NF-?B p65,p-NF-?B p65,COX-2 and iNOS were increased in the CIA model group,and the protein expression of PPAR-? was decreased(P<0.01).The protein expression of NF-?B p65,iNOS,and COX-2 decreased in each dose group of Shaogan Fuzi Decoction and Tripterygium glycoside tablet group,and PPAR-y protein expression increased,and the expression of p-NF?B p65 protein decreased in the high-dose Shaogan Fuzi Decoction group and Tripterygium glucoside tablet group compared with the model group(P<0.05).Conclusion:Shaogan Fuzi Decoction can reduce the levels of serum PGE2,NO,TNF-?,IL-1?,IL-6 in CIA model rats,and down-regulate the expression of NF-?B p65,p-NF-?B p65,COX-2,iNOS protein,and up-regulate PPAR-y protein expression,which indicates the regulatory effect of the prescription on the PPAR-?/NF-?B signaling pathway.The above research results suggest that Shaogan Fuzi Decoction can improve joint swelling and pathological damage in CIA rats,and its mechanism may be related to the regulation of PPAR-?/NF-?B signaling pathway and related inflammatory factors levels. |
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